This career development plan will prepare Dr. Irfan Lodhi to transition to a position as an independent academic investigator in metabolic research. The mentored phase (K99) of this 5 year project will be completed in the laboratory of Dr. Clay Semenkovich at Washington University School of Medicine. The overall objective of this project is to understand the role of lipogenic pathways in the activation of PPARg, a key regulator of adipose tissue development and metabolism. PPARg belongs to the peroxisome proliferator- activated receptor (PPAR) family of ligand-activated nuclear receptors, originally identified based on their ability to be activated by agents that promote peroxisome proliferation. Endogenous pathways of PPARg activation have remained unclear. Previous work suggests that de novo lipogenesis mediated by fatty acid synthase (FAS) is important for endogenous activation of PPARg in adipocytes. A mass spectrometry-based approach identified several FAS-dependent complex lipids as putative endogenous ligands for PPARg. These lipids are made in peroxisomes and require a previously unidentified peroxisomal enzyme named PexRAP for their synthesis. Knockdown of PexRAP in cultured cells impaired PPARg activation and adipogenesis, both of which could be rescued with rosiglitazone, a pharmacological activator of PPARg. Antisense oligonucleotide- mediated knockdown of PexRAP in mice decreased expression of PPARg-dependent genes, reduced fat mass, increased leanness, and improved insulin sensitivity. Additional studies suggested that genes involved in peroxisomal biogenesis increase during adipogenesis and are regulated by PPARg. Together, these studies led to the hypothesis that peroxisome-derived lipids activate PPARg and, reciprocally, PPARg promotes peroxisomal biogenesis, resulting in a feed-forward cycle that drives adipocyte differentiation. The following specific aims are designed to test this hypothesis. Aim 1 will determine if mice with adipose-specific deletion of PexRAP are protected from diet-induced and genetic forms of obesity and diabetes. Aim 2 will characterize the molecular mechanism of PexRAP function in adipocytes. Finally, Aim 3 will determine the role of peroxisomal biogenesis in adipocyte differentiation.